Based on the cause of decline in insulin activity, diabetes is generally classified into type I diabetes (juvenile-onset diabetes) and type II diabetes.
Type I diabetes is caused by autoimmune abnormalities in which pancreatic β cells secreting insulin are specifically destroyed (see Atkinson M A, Maclaren N K., N. Engl. J. Med. 331:1428-1436, 1994). To completely treat type I diabetes, transplantation which is one of the treatments for regenerating/replacing pancreatic β cells is considered. Such transplantations include pancreas transplantation and pancreatic islet transplantation. The purposes of these two kinds of transplantation are to enable the extremely exact regulation of blood glucose, thereby preventing hypoglycemia and long-term complication from being caused. It is not the sole purpose to release patients from daily bother in insulin treatment and improve the quality of life (QOL). As a means of achieving the goal to completely treat insulin-dependent diabetes, transplantation therapy has much greater potential than insulin treatment does. However, the pancreas transplantation has problems that the operative attack is serious, and that the complications caused by concomitantly transplanted exocrine glands may be severe. In contrast thereto, the purpose of pancreatic islet transplantation is to isolate and transplant pancreatic islet β cells by excising the exocrine glands which are the cause of complications associated with transplantation operation. Presently, the pancreatic islet transplantation is a promising method for the treatment of diabetes whereby the condition of a patient is brought back to precritical condition of diabetes by the most physiological procedure. In the case of type II diabetes wherein the secretion of insulin is partially maintained, insulin shortage is also caused by hyperglycemia-induced toxicity or β cell exhaustion when the stage of disease progresses. However, pancreatic islet transplantation has not been applied for type II diabetes on the basis of the reality that pancreatic islet for transplantation is in short supply. In the future, if a great number of pancreatic islets for transplantation can be supplied, there is a great possibility that pancreatic islet transplantation is indicated for insulin-dependent diabetes with insulin resistance.
In 2000, seven cases of clinical pancreatic islet transplantation were reported from University of Alberta in Edmonton, Canada (see Shapiro A M, Lakey J R, Ryan E A, et al., N. Engl. J. Med. 343:.230-238, 2000). The report described that by a novel method for using immunosuppressant called “Edmonton Protocol” later, all the cases with type I diabetes who underwent pancreatic islet transplantation became free from administration of insulin. At the present time, pancreatic islet transplantation is the closest to an ideal treatment method for patients who suffer from insulin-dependent diabetes.
Pancreatic islet is an endocrine gland tissue and its volume is at most 2% of the whole pancreas volume. Pancreatic islet is an aggregate of endocrine cells and comprises α cells, β cells, PP cells and δ cells and the like. Insulin which is the only endogenous hormone having the effect of decreasing blood sugar, is secreted by the β cells in pancreatic islet. The β cells account for 80 to 85% of the whole cells constructing pancreatic islet. The β cells not only secret insulin but also are capable of detecting sugars in blood. The aim of pancreatic islet transplantation is to replace and regenerate a system of decreasing blood sugar which once has declined, by isolating pancreatic islet from pancreas and transplanting it to a patient who suffers from insulin-dependent diabetes.
However, in the case of pancreatic islet transplantation at the present time, there are problems of safety caused by using immunosuppressant and further serious problem, shortage of pancreatic islets for transplantation. Even though the current pancreatic islet isolating technique were improve, the number of pancreas taken from neomorts for pancreas transplantation/isolation of pancreatic islet would be much smaller than the number of patients in need thereof. Therefore, there is no prospect of overcoming the shortage problem of pancreatic islets for transplantation.
Accordingly, manufacturing cells having functions comparable to those of pancreatic islet or pancreatic islet β cells provides large contribution to society and great impact on medical economy. Besides, in stem cell research progressing with great speed in these years and attracting public attention, possibility of differentiation induction toward a pancreatic endocrine cell is indicated (see Lumelsky N, Blondel O, Laeng P, et al., Science 292: 1389-1394, 2001; Assady S, Maor G, Amit M, et al., Diabetes 50: 1691-1697, 2001), which triggers further facilitation of an interest for producing an artificial pancreatic islet β cell.
As a source of cells replacing human mature pancreatic islet β cells, human ES cells and tissue stem cells, for example, are being intensively studied at the present time. Although it was reported that insulin expression was observed by differentiation inducing in some cells (murine ES cell and hepatic stem cell), it is still unclear which gene should be transferred at which stage for secreting insulin effectively. In addition, the use of such stem cell essentially involves the difficulty in control which arises from the fact that the stem cell has pluripotency and active proliferation potency. It is considered that plenty of time is necessary to put the cells to practical use hereafter.
Study using porcine tissue/cell progresses, whereas these problems of zoonotic infection, tissue compatibility and ethics have surfaced. In particular, potential risk connected with virus has become a serious problem. For example, there is a disease-producing risk that a porcine virus contained in a porcine organ or cells infects recipient (especially, it is impossible to eliminate a porcine endogenous retrovirus (PERV), because PERV is integrated in a chromosome), and there is a novel viral infection-spreading risk that the infection spreads to its family and medical staff, and further to society.
At the present time, therefore, establishment of a tractable insulin secreting human pancreatic islet cell line which is a source of cells replacing human mature pancreatic islet β cells is desired. Until now, many researchers have made aggressive efforts to immortalize a human pancreatic islet cell, but any human pancreatic islet cell line producing insulin has not been reported at all. Possible explanations are as follows; 1) gene transfer is difficult because insulin producing β cells exist in the inside of pancreatic islet, 2) cell-life extension is taken place but the complete immortalization of cells cannot be achieved because a tumor gene originated from a virus (such as a simian virus 40 tumor antigen) is used to establish an immortalized cell line. In fact, the frequency that human cells proliferate without limitation beyond proliferation decay is low (see Shay J W, Wright W E, Exp. Cell Res. 184: 109-118, 1989; Ray F A, Kraemer P M, Carcinogenesis 14: 1511-1516, 1993). Furthermore, it is reported that a probability of natural immortalization in a cell using SV40T in vitro is about 3.3×10−7; it is said that the cell further needs natural expression of autogenic telomerase activity (see, Bondar A G, Science 279: 349-352, 1998; Zhu J, et al., Proc. Natl. Acad. Sci. USA 96: 3723-3728, 1999; Halvorsen T L, et al., Mol. Cell. Biol. 19: 1864-1870, 1999).
An object of the present invention is to solve the problems in the prior art, and to provide a human pancreatic islet cell line capable of producing insulin and enabling easy obtainment of the number of cells which meets the demand.